The experimental approaches outlined in this proposal are designed to investigate molecular events regulating replication of the alphavirus Semliki Forest virus (SFV). We intend to study the regulation of replication of two full length RNAs of different polarity, a complementary copy of the infectious virion RNA and the progeny virion RNA. Synthesis of only the complementary RNA ceases early in infection. For these studies we will utilize certain temperature-sensitive mutants of SFV to determine if any alteration in the regulation of complementray RNA synthesis occurs. We intend to characterize the mode and regulation of transcription of a smaller than genome size RNA that is identical to the 3' third of the genome RNA. The sensitivity to ultraviolet irradiation and protection of the 5' terminal cap after annealing to various parts of the complementary RNA template will be identified. We will determine if synthesis of this smaller RNA involves internal initiation or a looping-out of part of the template RNA.